The Rev transactivator of human immunodeficiency virus type 1 (HIV-1) is a posttranscriptional regulatory protein that is essential for replication of the virus. Localized within the nuclei of infected cells, Rev acts through a poorly understood mechanism to allow cytoplasmic accumulation of certain incompletely spliced HIV-1 mRNAs that would otherwise remain sequestered in the nucleus. Because these mRNAs code for the major viral structural proteins, HIV- 1 proviruses that lack a functional rev gene are unable to produce new infectious virions. Rev is therefore of interest not only because of its unprecedented regulatory effect, but also as a promising target for antiretroviral therapy. Using techniques of molecular genetics, we are investigating the functional architecture and mechanism of action of Rev, searching for potential means of inhibiting its activity. Mutations are introduced systematically into Rev and its cis-acting target element in the viral genome, and the effects of these mutations are then characterized using a transient transfection assay. Our studies have identified, and will now further characterize, three critical peptide domains that mediate interactions of Rev monomers with each other, with putative cellular cofactors, and with the target RNAs. Essential features of each region will be mapped in detail to provide a basis for rational design of inhibitors, and we will search for equivalent domains in Rev-like proteins from other retroviruses and in selected cellular proteins. We will also pursue the observation that modified forms of these regions can block activity of wild-type Rev, as a possible approach to gene therapy. Using Rev fusion proteins that have altered target specificity, we will dissect the minimal requirements for this pathway of viral transactivation in vivo. We will also investigate how changes in the physiologic state of an infected T lymphocyte can modulate Rev activity. The information to be gained in these studies will be invaluable in designing novel pharmacologic or genetic strategies for inhibiting Rev, which could be used to maintain HIV-1 latency and slow the progression of disease in infected individuals.